scholarly journals The whole is greater than the sum of its parts: at the intersection of order, disorder, and kinetochore function

2020 ◽  
Vol 64 (2) ◽  
pp. 349-358
Author(s):  
Margaux R. Audett ◽  
Thomas J. Maresca
Keyword(s):  
Molecular Biology ◽  
Structure Function ◽  
Spatial Organization ◽  
Disordered Proteins ◽  
Intrinsically Disordered ◽  
The Past ◽  
Massive Structure ◽  
In Vitro Reconstitution ◽  
Kinetochore Function

Abstract The kinetochore (KT) field has matured tremendously since Earnshaw first identified CENP-A, CENP-B, and CENP-C [1,2]. In the past 35 years, the accumulation of knowledge has included: defining the parts list, identifying epistatic networks of interdependence within the parts list, understanding the spatial organization of subcomplexes into a massive structure – hundreds of megadaltons in size, and dissecting the functions of the KT in its entirety as well as of its individual parts. Like nearly all cell and molecular biology fields, the structure–function paradigm has been foundational to advances in the KT field. A point nicely highlighted by the fact that we are at the precipice of the in vitro reconstitution of a functional KT holo complex. Yet conventional notions of structure cannot provide a complete picture of the KT especially since it contains an abundance of unstructured or intrinsically disordered constituents. The combination of structured and disordered proteins within the KT results in an assembled system that is functionally greater than the sum of its parts.

scholarly journals Simple biophysics underpins collective conformations of the intrinsically disordered proteins of the Nuclear Pore Complex

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Andrei Vovk ◽  
Chad Gu ◽  
Michael G Opferman ◽  
Larisa E Kapinos ◽  
Roderick YH Lim ◽  
...  
Keyword(s):  
Spatial Organization ◽  
Intrinsically Disordered Proteins ◽  
Nucleocytoplasmic Transport ◽  
Transport Proteins ◽  
Disordered Proteins ◽  
Biophysical Model ◽  
Nuclear Pore ◽  
Intrinsically Disordered

Nuclear Pore Complexes (NPCs) are key cellular transporter that control nucleocytoplasmic transport in eukaryotic cells, but its transport mechanism is still not understood. The centerpiece of NPC transport is the assembly of intrinsically disordered polypeptides, known as FG nucleoporins, lining its passageway. Their conformations and collective dynamics during transport are difficult to assess in vivo. In vitro investigations provide partially conflicting results, lending support to different models of transport, which invoke various conformational transitions of the FG nucleoporins induced by the cargo-carrying transport proteins. We show that the spatial organization of FG nucleoporin assemblies with the transport proteins can be understood within a first principles biophysical model with a minimal number of key physical variables, such as the average protein interaction strengths and spatial densities. These results address some of the outstanding controversies and suggest how molecularly divergent NPCs in different species can perform essentially the same function.

scholarly journals Baicalein inhibits heparin-induced Tau aggregation by initializing non-toxic Tau oligomer formation

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Shweta Kishor Sonawane ◽  
Vladimir N. Uversky ◽  
Subashchandrabose Chinnathambi
Keyword(s):  
Intrinsically Disordered Proteins ◽  
Senile Plaques ◽  
Neuroblastoma Cells ◽  
Memory Loss ◽  
Disordered Proteins ◽  
Protein Accumulation ◽  
Scutellaria Baicalensis Georgi ◽  
Intrinsically Disordered ◽  
Tau Aggregation

Abstract Background Amyloid aggregate deposition is the key feature of Alzheimer’s disease. The proteinaceous aggregates found in the afflicted brain are the intra-neuronal neurofibrillary tangles formed by the microtubule-associated protein Tau and extracellular deposits, senile plaques, of amyloid beta (Aβ) peptide proteolytically derived from the amyloid precursor protein. Accumulation of these aggregates has manifestations in the later stages of the disease, such as memory loss and cognitive inabilities originating from the neuronal dysfunction, neurodegeneration, and brain atrophy. Treatment of this disease at the late stages is difficult, and many clinical trials have failed. Hence, the goal is to find means capable of preventing the aggregation of these intrinsically disordered proteins by inhibiting the early stages of their pathological transformations. Polyphenols are known to be neuroprotective agents with the noticeable potential against many neurodegenerative diseases, such as Alzheimer’s, Parkinson’s, and Prion diseases. Methods We analyzed the capability of Baicalein to inhibit aggregation of human Tau protein by a multifactorial analysis that included several biophysical and biochemical techniques. Results The potency of Baicalein, a polyphenol from the Scutellaria baicalensis Georgi, against in vitro Tau aggregation and PHF dissolution has been screened and validated. ThS fluorescence assay revealed the potent inhibitory activity of Baicalein, whereas ANS revealed its mechanism of Tau inhibition viz. by oligomer capture and dissociation. In addition, Baicalein dissolved the preformed mature fibrils of Tau thereby possessing a dual target action. Tau oligomers formed by Baicalein were non-toxic to neuronal cells, highlighting its role as a potent molecule to be screened against AD. Conclusion In conclusion, Baicalein inhibits aggregation of hTau40 by enhancing the formation of SDS-stable oligomers and preventing fibril formation. Baicalein-induced oligomers do not affect the viability of the neuroblastoma cells. Therefore, Baicalein can be considered as a lead molecule against Tau pathology in AD.

scholarly journals The Mysterious Unfoldome: Structureless, Underappreciated, Yet Vital Part of Any Given Proteome

2010 ◽  
Vol 2010 ◽  
pp. 1-14 ◽  
Author(s):  
Vladimir N. Uversky
Keyword(s):  
Large Scale ◽  
Intrinsically Disordered Proteins ◽  
Biologically Active ◽  
Disordered Proteins ◽  
Unfolded Proteins ◽  
Intrinsically Disordered ◽  
Proteome Level ◽  
Natively Unfolded ◽  
Natively Unfolded Proteins

Contrarily to the general believe, many biologically active proteins lack stable tertiary and/or secondary structure under physiological conditions in vitro. These intrinsically disordered proteins (IDPs) are highly abundant in nature and many of them are associated with various human diseases. The functional repertoire of IDPs complements the functions of ordered proteins. Since IDPs constitute a significant portion of any given proteome, they can be combined in an unfoldome; which is a portion of the proteome including all IDPs (also known as natively unfolded proteins, therefore, unfoldome), and describing their functions, structures, interactions, evolution, and so forth. Amino acid sequence and compositions of IDPs are very different from those of ordered proteins, making possible reliable identification of IDPs at the proteome level by various computational means. Furthermore, IDPs possess a number of unique structural properties and are characterized by a peculiar conformational behavior, including their high stability against low pH and high temperature and their structural indifference toward the unfolding by strong denaturants. These peculiarities were shown to be useful for elaboration of the experimental techniques for the large-scale identification of IDPs in various organisms. Some of the computational and experimental tools for the unfoldome discovery are discussed in this review.

Topological frustration leading to backtracking in a coupled folding-binding process

2021 ◽  
Author(s):  
Meng Gao ◽  
Ping Li ◽  
Zhengding Su ◽  
Yongqi Huang
Keyword(s):  
Structure Function ◽  
Intrinsically Disordered Proteins ◽  
Disordered Proteins ◽  
Biological Processes ◽  
Sequence Structure ◽  
Binding Process ◽  
Intrinsically Disordered

Intrinsically disordered proteins (IDPs) are abundant in all species. Their discovery challenges the traditional “sequence−structure−function” paradigm of protein science, because IDPs play important roles in various biological processes without preformed...

scholarly journals Contrasting Function of Structured N-Terminal and Unstructured C-Terminal Segments of Mycobacterium tuberculosis PPE37 Protein

mBio ◽  
2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Javeed Ahmad ◽  
Aisha Farhana ◽  
Rita Pancsa ◽  
Simran Kaur Arora ◽  
Alagiri Srinivasan ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Mycobacterium Tuberculosis ◽  
Host Cell ◽  
Terminal Segment ◽  
Host Protein ◽  
Host Cells ◽  
Productive Infection ◽  
Disordered Proteins ◽  
Intrinsically Disordered

ABSTRACT Pathogens frequently employ eukaryotic linear motif (ELM)-rich intrinsically disordered proteins (IDPs) to perturb and hijack host cell networks for a productive infection. Mycobacterium tuberculosis has a relatively high percentage of IDPs in its proteome, the significance of which is not known. The Mycobacterium-specific PE-PPE protein family has several members with unusually high levels of structural disorder and disorder-promoting Ala/Gly residues. PPE37 protein, a member of this family, carries an N-terminal PPE domain capable of iron binding, two transmembrane domains, and a disordered C-terminal segment harboring ELMs and a eukaryotic nuclear localization signal (NLS). PPE37, expressed as a function of low iron stress, was cleaved by M. tuberculosis protease into N- and C-terminal segments. A recombinant N-terminal segment (P37N) caused proliferation and differentiation of monocytic THP-1 cells, into CD11c, DC-SIGN (dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin)-positive semimature dendritic cells exhibiting high interleukin-10 (IL-10) but negligible IL-12 and also low tumor necrosis factor alpha (TNF-α) secretion—an environment suitable for maintaining tolerogenic immune cells. The C-terminal segment entered the macrophage nucleus and induced caspase-3-dependent apoptosis of host cells. Mice immunized with recombinant PPE37FL and PPE37N evoked strong anti-inflammatory response, validating the in vitro immunostimulatory effect. Analysis of the IgG response of PPE37FL and PPE37N revealed significant immunoreactivities in different categories of TB patients, viz. pulmonary TB (PTB) and extrapulmonary TB (EPTB), vis-a-vis healthy controls. These results support the role of IDPs in performing contrasting activities to modulate the host processes, possibly through molecular mimicry and cross talk in two spatially distinct host environments which may likely aid M. tuberculosis survival and pathogenesis. IMPORTANCE To hijack the human host cell machinery to enable survival inside macrophages, the pathogen Mycobacterium tuberculosis requires a repertoire of proteins that can mimic host protein function and modulate host cell machinery. Here, we have shown how a single protein can play multiple functions and hijack the host cell for the benefit of the pathogen. Full-length membrane-anchored PPE37 protein is cleaved into N- and C-terminal domains under iron-depleted conditions. The N-terminal domain facilitates the propathogen semimature tolerogenic state of dendritic cells, whereas the C-terminal segment is localized into host cell nucleus and induces apoptosis. The immune implications of these in vitro observations were assessed and validated in mice and also human TB patients. This study presents novel mechanistic insight adopted by M. tuberculosis to survive inside host cells.

scholarly journals Enhancing c-MYC degradation via 20S proteasome activation induces in vivo anti-tumor efficacy

2020 ◽  
Author(s):  
Evert Njomen ◽  
Theresa A. Lansdell ◽  
Allison Vanecek ◽  
Vanessa Benham ◽  
Matt P. Bernard ◽  
...  
Keyword(s):  
Therapeutic Strategy ◽  
Target Genes ◽  
Intrinsically Disordered Proteins ◽  
Therapeutic Potential ◽  
Human Cancer ◽  
Disordered Proteins ◽  
20S Proteasome ◽  
Intrinsically Disordered

SUMMARYEnhancing proteasome activity is a potential new therapeutic strategy to prevent the accumulation of aberrant high levels of protein that drive the pathogenesis of many diseases. Herein, we examine the use of small molecules to activate the 20S proteasome to reduce aberrant signaling by the undruggable oncoprotein c-MYC, to treat c-MYC driven oncogenesis. Overexpression of c-MYC is found in more than 50% of all human cancer but remains undruggable because of its highly dynamic intrinsically disordered 3-D conformation, which renders traditional therapeutic strategies largely ineffective. We demonstrate herein that small molecule activation of the 20S proteasome targets dysregulated intrinsically disordered proteins (IDPs), including c-MYC, and reduces cancer growth in vitro and in vivo models of multiple myeloma, and is even effective in bortezomib resistant cells and unresponsive patient samples. Genomic analysis of various cancer pathways showed that proteasome activation results in downregulation of many c-MYC target genes. Moreover, proteasome enhancement was well tolerated in mice and dogs. These data support the therapeutic potential of 20S proteasome activation in targeting IDP-driven proteotoxic disorders, including cancer, and demonstrate that this new therapeutic strategy is well tolerated in vivo.

scholarly journals Identifying sequence perturbations to an intrinsically disordered protein that determine its phase-separation behavior

2020 ◽  
Vol 117 (21) ◽  
pp. 11421-11431 ◽  
Author(s):  
Benjamin S. Schuster ◽  
Gregory L. Dignon ◽  
Wai Shing Tang ◽  
Fleurie M. Kelley ◽  
Aishwarya Kanchi Ranganath ◽  
...  
Keyword(s):  
Phase Separation ◽  
Intrinsically Disordered Proteins ◽  
Lipid Membrane ◽  
Coarse Grained ◽  
Disordered Proteins ◽  
Sequence Features ◽  
Intrinsically Disordered ◽  
Arginine Residues

Phase separation of intrinsically disordered proteins (IDPs) commonly underlies the formation of membraneless organelles, which compartmentalize molecules intracellularly in the absence of a lipid membrane. Identifying the protein sequence features responsible for IDP phase separation is critical for understanding physiological roles and pathological consequences of biomolecular condensation, as well as for harnessing phase separation for applications in bioinspired materials design. To expand our knowledge of sequence determinants of IDP phase separation, we characterized variants of the intrinsically disordered RGG domain from LAF-1, a model protein involved in phase separation and a key component of P granules. Based on a predictive coarse-grained IDP model, we identified a region of the RGG domain that has high contact probability and is highly conserved between species; deletion of this region significantly disrupts phase separation in vitro and in vivo. We determined the effects of charge patterning on phase behavior through sequence shuffling. We designed sequences with significantly increased phase separation propensity by shuffling the wild-type sequence, which contains well-mixed charged residues, to increase charge segregation. This result indicates the natural sequence is under negative selection to moderate this mode of interaction. We measured the contributions of tyrosine and arginine residues to phase separation experimentally through mutagenesis studies and computationally through direct interrogation of different modes of interaction using all-atom simulations. Finally, we show that despite these sequence perturbations, the RGG-derived condensates remain liquid-like. Together, these studies advance our fundamental understanding of key biophysical principles and sequence features important to phase separation.

scholarly journals Interplay of Structural Disorder and Short Binding Elements in the Cellular Chaperone Function of Plant Dehydrin ERD14

Cells ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 1856
Author(s):  
Nikoletta Murvai ◽  
Lajos Kalmar ◽  
Bianka Szalaine Agoston ◽  
Beata Szabo ◽  
Agnes Tantos ◽  
...  
Keyword(s):  
Heat Stress ◽  
Intrinsically Disordered Proteins ◽  
Stress Protein ◽  
Structural Disorder ◽  
Disordered Proteins ◽  
Sequence Motifs ◽  
E Coli ◽  
Intrinsically Disordered

Details of the functional mechanisms of intrinsically disordered proteins (IDPs) in living cells is an area not frequently investigated. Here, we dissect the molecular mechanism of action of an IDP in cells by detailed structural analyses based on an in-cell nuclear magnetic resonance experiment. We show that the ID stress protein (IDSP) A. thaliana Early Response to Dehydration (ERD14) is capable of protecting E. coli cells under heat stress. The overexpression of ERD14 increases the viability of E. coli cells from 38.9% to 73.9% following heat stress (50 °C × 15 min). We also provide evidence that the protection is mainly achieved by protecting the proteome of the cells. In-cell NMR experiments performed in E. coli cells show that the protective activity is associated with a largely disordered structural state with conserved, short sequence motifs (K- and H-segments), which transiently sample helical conformations in vitro and engage in partner binding in vivo. Other regions of the protein, such as its S segment and its regions linking and flanking the binding motifs, remain unbound and disordered in the cell. Our data suggest that the cellular function of ERD14 is compatible with its residual structural disorder in vivo.

scholarly journals Modelling the structure and interactions of intrinsically disordered peptides with multiple-replica, metadynamics-based sampling methods and force-field combinations

2021 ◽  
Author(s):  
Lunna Li ◽  
Tommaso Casalini ◽  
Paolo Arosio ◽  
Matteo Salvalaglio
Keyword(s):  
Force Field ◽  
Sampling Method ◽  
Intrinsically Disordered Proteins ◽  
Thermodynamic Characteristics ◽  
Building Blocks ◽  
Disordered Proteins ◽  
Intrinsically Disordered ◽  
Self Association ◽  
Disordered Peptides

Intrinsically disordered proteins (IDPs) play a key role in many biological processes, including the formation of biomolecular condensates within cells. A detailed characterization of their configurational ensemble and structure-function paradigm is crucial for understanding their biological activity and for exploiting them as building blocks in material sciences. In this work, we incorporate bias-exchange metadynamics and parallel-tempering well-tempered metadynamics with CHARMM36m and CHARMM22* to explore the structural and thermodynamic characteristics of a short archetypal disordered sequence derived from a DEAD-box protein. The conformational landscapes emerging from our simulations are largely congruent across methods and forcefields. Nevertheless, differences in fine details emerge from varying forcefield/sampling method combinations. For this protein, our analysis identifies features that help to explain the low propensity of this sequence to undergo self-association in vitro, which can be common to all force-field/sampling method combinations. Overall, our work demonstrates the importance of using multiple force-field/enhanced sampling method combinations for accurate structural and thermodynamic information in the study of general disordered proteins.

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